Staphylokinase in the supernatant of cultures of Staphylococcus aureus was concentrated and then purified twice successively by isoelectric focusing. The total activity of staphylokinase was increased about 100 per cent. after the first isoelectric focusing. The cause of this was investigated. It was also shown by means of this separation technique that staphylokinase is a heterogenous protein, with isoelectric points at pH 5.8 (component A), 6.2 (component B) and 6.8 (component C). The degree of purification was calculated.